Dictyostelium discoideum is a primitive eukaryotic microorganism commonly called a slime mold, or more specifically, a cellular slime mold. The name is derived from the two extreme states of the microorganism from a macroscopic perspective. When actively growing, the D. discoideum grow as single cell amoeba. At this stage they have no cell wall, hence their appearance as a thin film (or slime). Upon starvation on a solid medium, the independent cells aggregate to form a colony. The colony exhibits traits of a multicellular organism in that it migrates in the form called a slug and then differentiates, with the posterior cells of the slug forming a foot, the anterior cells forming a stalk and the middle cells forming a fruiting body. The organism is found naturally on the surface of soil and dung. The wild type amoeba obtains nutrients exclusively by ingestion (phagocytosis) of whole bacteria; for this reason they are sometimes referred to as carnivorous. Axenic mutants of D. discoideum have been isolated which are capable of growth without coculture of "food" bacteria and which therefore can be grown on soluble media. The present invention relates to a novel dipeptidylaminopeptidase isolated from D. discoideum.
Dipeptidylaminopeptidases (DAP) are enzymes which hydrolyze the penultimate amino terminal peptide bond releasing dipeptides from the unblocked amino-termini of peptides and proteins. There are currently four classes of dipeptidylaminopeptidases (designated DAP-I, DAP-II, DAP-III and DAP-IV) which are distinguished based on their physical characteristics and the rates at which they catalyze cleavage with various amino-terminal peptide sequences. DAP I is a relatively non-specific DAP that will catalyze the release of many dipeptide combinations from the unblocked amino termini of peptides and proteins. DAP I shows little or no activity if the emergent dipeptide is X-Pro, Arg-X, or Lys-X (where X is any amino acid). DAP II shows a preference for amino terminal dipeptide sequences that begin with Arg-X or Lys-X, and to a lesser extent, X-Pro. DAP-II exhibits significantly lower cleavage rates versus most other dipeptide combinations. DAP III appears to have a propensity toward amino terminal dipeptide sequences of the form Arg-Arg and Lys-Lys. DAP IV shows its highest rate of hydrolytic activity toward dipeptide sequences of the form X-Pro. The DAP enzymes, particularly DAP-I and DAP-IV, have been shown to be useful in processing proteins. The present invention concerns a novel DAP from Dictyostelium discoideum which is useful in processing recombinant proteins with an even numbered amino acid N-terminal extension.